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Importance of secreted bacterial RNA in bacterial-host interactions in the gut
Microbial Pathogenesis 104 (2017) 161e163
Contents lists available at ScienceDirect
Microbial Pathogenesis journal homepage: www.elsevier.com/locate/micpath
Importance of secreted bacterial RNA in bacterial-host interactions in the gut Anubrata Ghosal Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, United States
a r t i c l e i n f o
a b s t r a c t
Article history: Received 23 November 2016 Received in revised form 12 January 2017 Accepted 18 January 2017 Available online 19 January 2017
Bacteria are ubiquitous in nature and found to be associated with human. Humans are benefited significantly from these associated bacteria. Our understanding is quite limited about these beneficial associations and how bacteria communicate with the host. It is assumed that secreted bacterial products contribute in bacterial-host signaling. A few studies have shown that bacteria secrete RNA into their extracellular medium, and these secreted RNA are capable of altering the host immune response. Multiple studies in eukaryotes have confirmed that secreted RNA can influence the functioning of other cells and their role in the development of several diseases, although not much is known about the composition of secreted bacterial RNA, how they are trafficked and how they impact on the functioning of host cells. By uncovering the beneficial role of secreted bacterial RNA, it would be possible to improve the human health. © 2017 Elsevier Ltd. All rights reserved.
Keywords: Secreted bacterial RNA Bacterial-host interaction and gut
Contents References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 162
In nature, bacteria are found to be associated with a range of eukaryotes spanning from single-celled organisms, e.g. protists [1], to higher organisms, e.g. animals including human. The number of microorganisms living in the human body is more than the total number of cells present in the human body [2]. These associated microorganisms (microbiome), which are comprised of bacteria, archaea and eukarya, are coevolved with human and have a mutualistic relationship with human. Gut is one of the major sources of microorganism in the human body. The number of genes represented by the gut microbiome is more than 100 times larger in size than the total number of genes present in human [3], and the function of these additional genes is believed to be tremendously important in maintaining people's health. For example, the gut microbiome participate in digesting complex polysaccharide diet and help in absorption [3]. They may involve in shaping host immune response, preventing colonization of pathogenic microorganisms and many more unknown functions
E-mail address: [email protected]. http://dx.doi.org/10.1016/j.micpath.2017.01.032 0882-4010/© 2017 Elsevier Ltd. All rights reserved.
[4]. The majority of the gut microbiome are represented by the bacteria [3] and located in the distal part of the gut, colon [2]. Only two bacterial divisions, Bacteroidetes and Firmicutes, among 70 known bacterial divisions, represent 90% of the total bacteria present in the colon [2]. The bacterial density was recorded highest in the human gut in comparison to any known ecosystems [4,5]. Interestingly, bacteria are found to be more diverse at different locations in the gut than among individuals [6]. We have just begun to understand the microbiome's role in human health and disease. There is a lot to discover. One of the interesting areas of research is the role of secreted bacterial products in bacterial-host signaling. It is known that bacteria release cellular products into its surrounding medium, and, in bacteria, secretion occurs through the secretory systems and/or by the release of outer membrane vesicles (OMVs) [5,6]. Several molecules such as proteins, DNA, RNA, small autoinducer, toxins, enzymes and glycoproteins are known to be released by bacteria [6e9]. It is significantly challenging to characterize products released by the gut bacteria and also to analyze their impact on human cells as
162
A. Ghosal / Microbial Pathogenesis 104 (2017) 161e163
most of the gut bacteria are facultative to strict anaerobes and not all are culturable in artificial medium [10]. Recently, a microfluidic gut-on-chip system was presented allowing co-culture of gut bacteria with human cells and isolating products released by bacteria in real-time [10]. There is a considerable gap in our understanding of how secreted bacterial products, especially RNA, influence the host cell functions, but an array of studies have reported how human cells identify bacterial products and characterized their impact on immune cells. The interaction between the pathogen-associated molecular patterns (PAMPs) and the pattern recognition receptors (PRRs) such as Toll-like receptors (TLRs) and NOD-like receptors (NLRs) is important for the recognition of bacterial components [11]. PAMPs are comprised of conserved molecular motifs of exogenous origin, including peptidoglycans, lipoproteins, lipopolysaccharides (LPS), flagellins, CpG DNAs and RNAs of bacterial origin [12e14]. Study on Moraxella catarrhalis showed that OMV-borne DNA are recognized by the host TLR9 receptor inside the endosome of human B cells, and trigger secretion of IL6 and IgM [15]. Further, a study on Bartonella henselae demonstrated that type-IV secretory system is involved in the transfer of DNA into the human cells [16]. On the other hand, several studies have already indicated that mammalian cells are capable of distinguishing between endogenous and bacterial RNAs [13,17e19], and cytosolic receptors such as RIG-I, MDA5 and STING are involved in the recognition of bacterial RNAs [13,14]. Further, the bacterial RNA are known to stimulate the activation of caspase-1 and production of proinflammatory cytokines IL-18 and IL-1b; Cryopyrin, which is one of the key components of inflammasome, play an important role in host defence activated by bacterial RNA [13]. tRNA, mRNA, and rRNA originating from bacteria are capable of activating NLRP3 inflammasome in human macrophages [20]. Additionally, it is known that Toll-like receptors (TLR) such as TLR1 [21] and TLR8 [22] can sense miRNA. Therefore, it would be interesting to investigate TLR1 and TLR8 roles in detecting secreted bacterial RNA. Furthermore, a study showed that the bacterial secreted RNA are necessary for generating innate immune response against Listeria infection, and murine macrophages release IFNb in response to secreted RNA [14]. Similarly, another study highlighted the role of secreted Mycobacterium RNA in inducing pathogenesis and characterized secreted RNA as an early apoptotic factor. These low molecular weight secreted RNA are capable of inducing apoptosis via caspase-8 dependent pathway [23]. Both these studies lack a detailed characterization of secreted RNA. Very recently, a study on OMVs derived from Pseudomonas aeruginosa showed that OMV associated small RNA are actually able to suppress immune response in the host. They specifically characterized a methionine tRNA fragment, which transports to host cell via OMV and participates in the alteration of host gene expression, leading to a reduction of cytokine release [24]. Uropathogenic Escherichia coli (UPEC) strain 536 also releases RNA of different sizes, which also transported to human cells via OMVs [25]. Furthermore, the interkingdom crosstalk has shown to be exist between fungi and plant, where fungal small RNA by binding to Argonaute 1 protein is able to selectively silence the function of the host genes, but, in this study, they did not show how these fungi small RNA are transported to the plant cells [26]. E. coli, Porphyromonas gingivalis and Vibrio cholerae O1 El Tor strain are known to release RNA in their extracellular environment [9,27,28]. Analysis of the role of these secreted RNA in cross-kingdom signaling may uncover pathways that are essential for either sensing and/or combating bacterial infections. Secretion of RNA has also been observed in marine photosynthetic bacterium Rhodovulum sulfidophilu, and analysis showed that tRNA and rRNA represent the majority of the secreted RNA [29].
In the gut, bacteria initially encounter the host epithelial cells. It is widely known that innate immune cells, especially the dendritic cells, are involved in sampling of foreign materials present in the lumen of human gut [4]. Once bacteria cross the host epithelial barrier, they are either phagocytosed by the resident macrophages in the lamina propria or engulfed by the dendritic cells which later interact with the B and T cells and activate them. These activated B and T cells circulate via blood and lymph, and reach to the lamina propria, where they secrete effector molecules [30]. Together with the immune cells, epithelial cells play a major role in adjusting bacterial load on the lumen side of the gut, and molecules secreted by the epithelial cells also participate in differentiation and activation of immune cells in the gut [31]. It would be important to study how bacterial released RNA alters the secretory profile of host cells. Further, the uptake of secreted bacterial RNA into the host cells can vary depend on their association with other bacterial components. A couple of studies have shown that how OMVs release their content into the host cells. OMVs originating from Pseudomonas aerugina first fuse with lipid rafts and then release their content into the host cytoplasm following an actin-dependent pathway [32]. Moraxella catarrhalis OMVs, which they use to divert the host humoral response, internalize into the B cells following crosslinking with IgD on the surface of host cells [15]. Except for the OMV-mediated transport of bacterial secreted RNA, nothing is known about how the RNA-protein or lipid complexes are taken up by the host cells. A signification portion of secreted bacterial RNA is not associated with OMVs [9]. To my best knowledge, no bacterial secretory system has been identified that involve in the release of RNA. Identification of molecules that participate in the transportation of bacterial secreted RNA will help in designing vehicles for delivering cargo into the host cells. It could be that the secretory bacterial RNA are sensed by the host receptors, but in the cell they may function as antisense molecules. Inside the host cell, these secreted bacterial RNA may remain stable due to their association with proteins and/or lipids. RNA modifications could also be important for their stability. Further, it may be that the sequence motifs, modifications and/or the secondary structure of bacterial extracellular RNAs are important for their recognition as exogenous RNA by the host cells. Furthermore, bacterial secreted RNA, which are capable of inducing immune response, can be used as adjuvants to boost immune system. A study showed that specific RNA molecules are recognized by human TLR8 receptor and able to generate immune response [33]. Moreover, bacterial secreted RNA can be used as biomarker to detect the presence of specific bacteria in the gut. Overall, by studying bacterial secreted RNA, we will able to understand the part of complex interactions that exist between bacteria and humans in the gut and design strategies to improve human health. References [1] E.C.M. Nowack, M. Melkonian, Endosymbiotic associations within protists, Philos. Trans. R. Soc. Lond. B. Biol. Sci. 365 (2010) 699e712. [2] P.J. Turnbaugh, R.E. Ley, M. Hamady, C.M. Fraser-Liggett, R. Knight, J.I. Gordon, The human microbiome project, Nature 449 (2007) 804e810. [3] F. B€ ackhed, R.E. Ley, J.L. Sonnenburg, D. a Peterson, J.I. Gordon, Host-bacterial mutualism in the human intestine, Science 307 (2005) 1915e1920. [4] L.V. Hooper, D.R. Littman, A.J. Macpherson, Interactions between the microbiota and the immune system, Science 336 (2012) 1268e1273. [5] T.-T. Tseng, B.M. Tyler, J.C. Setubal, Protein secretion systems in bacterial-host associations, and their description in the Gene Ontology, BMC Microbiol. 9 (2009) 1471e2180. [6] Y. Shen, M.L. Giardino Torchia, G.W. Lawson, C.L. Karp, J.D. Ashwell, S.K. Mazmanian, Outer membrane vesicles of a human commensal mediate immune regulation and disease protection, Cell Host Microbe 12 (2012) 509e520. [7] S. Molloy, Quorum sensing: setting the threshold, Nat. Rev. Microbiol. 8 (2010) 388e389.
A. Ghosal / Microbial Pathogenesis 104 (2017) 161e163 [8] C.M. Waters, B.L. Bassler, Quorum sensing: cell-to-cell communication in bacteria, Annu. Rev. Cell Dev. Biol. 21 (2005) 319e346. [9] A. Ghosal, B.B. Upadhyaya, J.V. Fritz, A. Heintz-Buschart, M.S. Desai, D. Yusuf, D. Huang, A. Baumuratov, K. Wang, D. Galas, P. Wilmes, The extracellular RNA complement of Escherichia coli, Microbiologyopen 4 (2015) 252e266. [10] P. Shah, J.V. Fritz, E. Glaab, M.S. Desai, K. Greenhalgh, A. Frachet, M. Niegowska, M. Estes, C. Jager, C. Seguin-Devaux, F. Zenhausern, P. Wilmes, A microfluidics-based in vitro model of the gastrointestinal human-microbe interface, Nat. Commun. 7 (2016). [11] C.N. Casson, A.M. Copenhaver, E.E. Zwack, H.T. Nguyen, T. Strowig, B. Javdan, W.P. Bradley, T.C. Fung, R.A. Flavell, I.E. Brodsky, S. Shin, Caspase-11 activation in response to bacterial secretion systems that access the host cytosol, PLoS Pathog. 9 (2013) e1003400. [12] K.H.G. Mills, TLR-dependent T cell activation in autoimmunity, Nat. Rev. Immunol. 11 (2011) 807e822. €ren, M. Body-Malapel, A. Amer, J.-H. Park, L. Franchi, [13] T.-D. Kanneganti, N. Ozo J. Whitfield, W. Barchet, M. Colonna, P. Vandenabeele, J. Bertin, A. Coyle, ~ ez, Bacterial RNA and small antiviral compounds E.P. Grant, S. Akira, G. Nún activate caspase-1 through cryopyrin/Nalp3, Nature 440 (2006) 233e236. €ttcher, T. Hain, [14] Z. Abdullah, M. Schlee, S. Roth, M.A. Mraheil, W. Barchet, J. Bo S. Geiger, Y. Hayakawa, J.H. Fritz, F. Civril, K.-P. Hopfner, C. Kurts, J. Ruland, G. Hartmann, T. Chakraborty, P.A. Knolle, RIG-I detects infection with live Listeria by sensing secreted bacterial nucleic acids, EMBO J. 31 (2012) 4153e4164. [15] M. Vidakovics, J. Jendholm, B cell activation by outer membrane vesiclesda novel virulence mechanism, PLoS Pathog. 6 (2010) e1000724. € der, R. Schuelein, M. Quebatte, C. Dehio, Conjugative DNA transfer [16] G. Schro into human cells by the VirB/VirD4 type IV secretion system of the bacterial pathogen Bartonella henselae, Proc. Natl. Acad. Sci. U. S. A. 108 (2011) 14643e14648. [17] M. Oldenburg, A. Krüger, R. Ferstl, A. Kaufmann, G. Nees, A. Sigmund, B. Bathke, H. Lauterbach, M. Suter, S. Dreher, U. Koedel, S. Akira, T. Kawai, J. Buer, H. Wagner, S. Bauer, H. Hochrein, C.J. Kirschning, TLR13 recognizes bacterial 23S rRNA devoid of erythromycin resistance-forming modification, Science 337 (2012) 1111e1115. [18] L.E. Sander, M.J. Davis, M.V. Boekschoten, D. Amsen, C.C. Dascher, B. Ryffel, J.A. Swanson, M. Müller, J.M. Blander, Detection of prokaryotic mRNA signifies microbial viability and promotes immunity, Nature 474 (2011) 385e389. , S. Xu, D. Weissman, P.A. Cohen, B.J. Czerniecki, Cutting [19] G.K. Koski, K. Kariko edge: innate immune system discriminates between RNA containing bacterial versus eukaryotic structural features that prime for high-level IL-12 secretion by dendritic cells, J. Immunol. 172 (2004) 3989e3993. [20] W. Sha, H. Mitoma, S. Hanabuchi, M. Bao, L. Weng, N. Sugimoto, Y. Liu, Z. Zhang, J. Zhong, B. Sun, Y.-J. Liu, Human NLRP3 inflammasome senses
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Contents lists available at ScienceDirect
Microbial Pathogenesis journal homepage: www.elsevier.com/locate/micpath
Importance of secreted bacterial RNA in bacterial-host interactions in the gut Anubrata Ghosal Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, United States
a r t i c l e i n f o
a b s t r a c t
Article history: Received 23 November 2016 Received in revised form 12 January 2017 Accepted 18 January 2017 Available online 19 January 2017
Bacteria are ubiquitous in nature and found to be associated with human. Humans are benefited significantly from these associated bacteria. Our understanding is quite limited about these beneficial associations and how bacteria communicate with the host. It is assumed that secreted bacterial products contribute in bacterial-host signaling. A few studies have shown that bacteria secrete RNA into their extracellular medium, and these secreted RNA are capable of altering the host immune response. Multiple studies in eukaryotes have confirmed that secreted RNA can influence the functioning of other cells and their role in the development of several diseases, although not much is known about the composition of secreted bacterial RNA, how they are trafficked and how they impact on the functioning of host cells. By uncovering the beneficial role of secreted bacterial RNA, it would be possible to improve the human health. © 2017 Elsevier Ltd. All rights reserved.
Keywords: Secreted bacterial RNA Bacterial-host interaction and gut
Contents References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 162
In nature, bacteria are found to be associated with a range of eukaryotes spanning from single-celled organisms, e.g. protists [1], to higher organisms, e.g. animals including human. The number of microorganisms living in the human body is more than the total number of cells present in the human body [2]. These associated microorganisms (microbiome), which are comprised of bacteria, archaea and eukarya, are coevolved with human and have a mutualistic relationship with human. Gut is one of the major sources of microorganism in the human body. The number of genes represented by the gut microbiome is more than 100 times larger in size than the total number of genes present in human [3], and the function of these additional genes is believed to be tremendously important in maintaining people's health. For example, the gut microbiome participate in digesting complex polysaccharide diet and help in absorption [3]. They may involve in shaping host immune response, preventing colonization of pathogenic microorganisms and many more unknown functions
E-mail address: [email protected]. http://dx.doi.org/10.1016/j.micpath.2017.01.032 0882-4010/© 2017 Elsevier Ltd. All rights reserved.
[4]. The majority of the gut microbiome are represented by the bacteria [3] and located in the distal part of the gut, colon [2]. Only two bacterial divisions, Bacteroidetes and Firmicutes, among 70 known bacterial divisions, represent 90% of the total bacteria present in the colon [2]. The bacterial density was recorded highest in the human gut in comparison to any known ecosystems [4,5]. Interestingly, bacteria are found to be more diverse at different locations in the gut than among individuals [6]. We have just begun to understand the microbiome's role in human health and disease. There is a lot to discover. One of the interesting areas of research is the role of secreted bacterial products in bacterial-host signaling. It is known that bacteria release cellular products into its surrounding medium, and, in bacteria, secretion occurs through the secretory systems and/or by the release of outer membrane vesicles (OMVs) [5,6]. Several molecules such as proteins, DNA, RNA, small autoinducer, toxins, enzymes and glycoproteins are known to be released by bacteria [6e9]. It is significantly challenging to characterize products released by the gut bacteria and also to analyze their impact on human cells as
162
A. Ghosal / Microbial Pathogenesis 104 (2017) 161e163
most of the gut bacteria are facultative to strict anaerobes and not all are culturable in artificial medium [10]. Recently, a microfluidic gut-on-chip system was presented allowing co-culture of gut bacteria with human cells and isolating products released by bacteria in real-time [10]. There is a considerable gap in our understanding of how secreted bacterial products, especially RNA, influence the host cell functions, but an array of studies have reported how human cells identify bacterial products and characterized their impact on immune cells. The interaction between the pathogen-associated molecular patterns (PAMPs) and the pattern recognition receptors (PRRs) such as Toll-like receptors (TLRs) and NOD-like receptors (NLRs) is important for the recognition of bacterial components [11]. PAMPs are comprised of conserved molecular motifs of exogenous origin, including peptidoglycans, lipoproteins, lipopolysaccharides (LPS), flagellins, CpG DNAs and RNAs of bacterial origin [12e14]. Study on Moraxella catarrhalis showed that OMV-borne DNA are recognized by the host TLR9 receptor inside the endosome of human B cells, and trigger secretion of IL6 and IgM [15]. Further, a study on Bartonella henselae demonstrated that type-IV secretory system is involved in the transfer of DNA into the human cells [16]. On the other hand, several studies have already indicated that mammalian cells are capable of distinguishing between endogenous and bacterial RNAs [13,17e19], and cytosolic receptors such as RIG-I, MDA5 and STING are involved in the recognition of bacterial RNAs [13,14]. Further, the bacterial RNA are known to stimulate the activation of caspase-1 and production of proinflammatory cytokines IL-18 and IL-1b; Cryopyrin, which is one of the key components of inflammasome, play an important role in host defence activated by bacterial RNA [13]. tRNA, mRNA, and rRNA originating from bacteria are capable of activating NLRP3 inflammasome in human macrophages [20]. Additionally, it is known that Toll-like receptors (TLR) such as TLR1 [21] and TLR8 [22] can sense miRNA. Therefore, it would be interesting to investigate TLR1 and TLR8 roles in detecting secreted bacterial RNA. Furthermore, a study showed that the bacterial secreted RNA are necessary for generating innate immune response against Listeria infection, and murine macrophages release IFNb in response to secreted RNA [14]. Similarly, another study highlighted the role of secreted Mycobacterium RNA in inducing pathogenesis and characterized secreted RNA as an early apoptotic factor. These low molecular weight secreted RNA are capable of inducing apoptosis via caspase-8 dependent pathway [23]. Both these studies lack a detailed characterization of secreted RNA. Very recently, a study on OMVs derived from Pseudomonas aeruginosa showed that OMV associated small RNA are actually able to suppress immune response in the host. They specifically characterized a methionine tRNA fragment, which transports to host cell via OMV and participates in the alteration of host gene expression, leading to a reduction of cytokine release [24]. Uropathogenic Escherichia coli (UPEC) strain 536 also releases RNA of different sizes, which also transported to human cells via OMVs [25]. Furthermore, the interkingdom crosstalk has shown to be exist between fungi and plant, where fungal small RNA by binding to Argonaute 1 protein is able to selectively silence the function of the host genes, but, in this study, they did not show how these fungi small RNA are transported to the plant cells [26]. E. coli, Porphyromonas gingivalis and Vibrio cholerae O1 El Tor strain are known to release RNA in their extracellular environment [9,27,28]. Analysis of the role of these secreted RNA in cross-kingdom signaling may uncover pathways that are essential for either sensing and/or combating bacterial infections. Secretion of RNA has also been observed in marine photosynthetic bacterium Rhodovulum sulfidophilu, and analysis showed that tRNA and rRNA represent the majority of the secreted RNA [29].
In the gut, bacteria initially encounter the host epithelial cells. It is widely known that innate immune cells, especially the dendritic cells, are involved in sampling of foreign materials present in the lumen of human gut [4]. Once bacteria cross the host epithelial barrier, they are either phagocytosed by the resident macrophages in the lamina propria or engulfed by the dendritic cells which later interact with the B and T cells and activate them. These activated B and T cells circulate via blood and lymph, and reach to the lamina propria, where they secrete effector molecules [30]. Together with the immune cells, epithelial cells play a major role in adjusting bacterial load on the lumen side of the gut, and molecules secreted by the epithelial cells also participate in differentiation and activation of immune cells in the gut [31]. It would be important to study how bacterial released RNA alters the secretory profile of host cells. Further, the uptake of secreted bacterial RNA into the host cells can vary depend on their association with other bacterial components. A couple of studies have shown that how OMVs release their content into the host cells. OMVs originating from Pseudomonas aerugina first fuse with lipid rafts and then release their content into the host cytoplasm following an actin-dependent pathway [32]. Moraxella catarrhalis OMVs, which they use to divert the host humoral response, internalize into the B cells following crosslinking with IgD on the surface of host cells [15]. Except for the OMV-mediated transport of bacterial secreted RNA, nothing is known about how the RNA-protein or lipid complexes are taken up by the host cells. A signification portion of secreted bacterial RNA is not associated with OMVs [9]. To my best knowledge, no bacterial secretory system has been identified that involve in the release of RNA. Identification of molecules that participate in the transportation of bacterial secreted RNA will help in designing vehicles for delivering cargo into the host cells. It could be that the secretory bacterial RNA are sensed by the host receptors, but in the cell they may function as antisense molecules. Inside the host cell, these secreted bacterial RNA may remain stable due to their association with proteins and/or lipids. RNA modifications could also be important for their stability. Further, it may be that the sequence motifs, modifications and/or the secondary structure of bacterial extracellular RNAs are important for their recognition as exogenous RNA by the host cells. Furthermore, bacterial secreted RNA, which are capable of inducing immune response, can be used as adjuvants to boost immune system. A study showed that specific RNA molecules are recognized by human TLR8 receptor and able to generate immune response [33]. Moreover, bacterial secreted RNA can be used as biomarker to detect the presence of specific bacteria in the gut. Overall, by studying bacterial secreted RNA, we will able to understand the part of complex interactions that exist between bacteria and humans in the gut and design strategies to improve human health. References [1] E.C.M. Nowack, M. Melkonian, Endosymbiotic associations within protists, Philos. Trans. R. Soc. Lond. B. Biol. Sci. 365 (2010) 699e712. [2] P.J. Turnbaugh, R.E. Ley, M. Hamady, C.M. Fraser-Liggett, R. Knight, J.I. Gordon, The human microbiome project, Nature 449 (2007) 804e810. [3] F. B€ ackhed, R.E. Ley, J.L. Sonnenburg, D. a Peterson, J.I. Gordon, Host-bacterial mutualism in the human intestine, Science 307 (2005) 1915e1920. [4] L.V. Hooper, D.R. Littman, A.J. Macpherson, Interactions between the microbiota and the immune system, Science 336 (2012) 1268e1273. [5] T.-T. Tseng, B.M. Tyler, J.C. Setubal, Protein secretion systems in bacterial-host associations, and their description in the Gene Ontology, BMC Microbiol. 9 (2009) 1471e2180. [6] Y. Shen, M.L. Giardino Torchia, G.W. Lawson, C.L. Karp, J.D. Ashwell, S.K. Mazmanian, Outer membrane vesicles of a human commensal mediate immune regulation and disease protection, Cell Host Microbe 12 (2012) 509e520. [7] S. Molloy, Quorum sensing: setting the threshold, Nat. Rev. Microbiol. 8 (2010) 388e389.
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